Compartments within cellular membranes, specifically regions with distinct protein and lipid compositions, have been implicated in critical membrane functions such as cellular signaling. These regions have also been implicated in pathogen and toxin entry, and in pathologies like Alzheimer's disease. In this project, we aim to investigate, at the single molecule level, the mechanism(s) by which receptor proteins are localized to specific regions in an Escherichia coli K- 12 model system, with simpler membrane architecture than what is found in eukaryotes. Individual fluorescently labeled serine chemoreceptors (Tsr) in the inner membrane of E. coli will be imaged at video frame rates up to 1000 frames per second to determine how the "Tsr membrane domain" is assembled and maintained. Single molecule imaging has the combined benefits of the ability to observe rare events that would be lost in bulk, multi-molecule imaging and the ability to localize molecules to positions much smaller than the Raleigh limit of resolution. As such, the dynamics of the molecules in their domain and their entrance and possible exchange with free molecules outside the domain can be accurately monitored. Imaging of Tsr will be carried out in normal and mutant cells that have been genetically altered in cellular processors that are predicted to affect localization in accordance with testable hypotheses (e. g., lipid composition, specific partner protein interactions, and the bacterial cytoskeleton). In these ways, we expect to gain a deeper understanding of molecular and physical mechanisms by which integral membrane proteins are targeted to specific locations or membrane domains. [unreadable] [unreadable] Project Narrative: All cells are surrounded by a membrane that contain a number of different molecules that are important for acquisition of nutrients, deposition of cell waste products, and a myriad of other interactions with its surroundings. These membrane molecules can be static or highly mobile depending upon their function. Understanding how cells control the dynamics of its membrane molecules is important for developing new strategies for combating processes like viral infection and preventing diseases such as Alzheimer's disease. New technological advances for high-speed single molecule detection developed in this project will also be directly applicable to many other important basic biological problems. [unreadable] [unreadable] [unreadable]